TOP ＞ Symposium

Symposium 8 Current topics in GABA synthetic enzyme and psychiatric disorder シンポジウム8 GABA合成酵素と精神疾患、最近の知見 SY8-1 Cortical GAD67 deficits and the pathophysiology of schizophrenia 統合失調症の大脳皮質回路変化とGAD67発現低下 Hashimoto Takanori（橋本 隆紀）1,2 1Department of Psychiatry and Behavioral Science, Kanazawa University 2Department of Psychiatry, University of Pittsburgh Multiple postmortem studies reported lower mRNA levels of the 67 kDa isoform of glutamic acid decarboxylase (GAD67), the enzyme responsible for cortical γ-aminobutyric acid (GABA) synthesis, in the dorsolateral prefrontal cortex (DLPFC) of schizophrenia subjects. GAD67 mRNA levels are markedly reduced in the subset of GABA neurons that express parvalbumin (PV) without a change in the number of GABA neurons or PV neurons. Consistently, GAD67 protein levels were found to be lower in the DLPFC, especially in the presynaptic terminals of PV neurons. As reduced GABA synthesis in PV neurons could cause a shift in cortical activity, we hypothesized that lower levels of other transcripts, including GABA transporter-1, PV, brain-derived neurotrophic factor (BDNF), tropomyosin receptor kinase B, somatostatin, GABAA receptor α1 subunit and KCNS3 potassium channel subunit mRNAs, in the DLPFC of schizophrenia subjects, are secondary to the GAD67 deficit in PV neurons. In order to test this hypothesis, we analyzed cortical levels of these transcripts in mice with a PV neuron-specific GAD67 knockout. Using in situ hybridization, we found that none of these transcripts exhibited a lower expression in the knockout mice. In contrast, PV, BDNF, KCNS3 and GAD65 mRNA levels were higher in the homozygous mice. In addition, our behavioral test battery failed to detect a change in sensorimotor gating or working memory, although the homozygous mice exhibited increased spontaneous activities. These findings indicate that reduced GAD67 expression in PV neurons is not an upstream cause of the downregulated expression of other transcripts or of the cognitive/behavioral impairments present in subjects with schizophrenia. SY8-2 GAD knockout rats for a model of psychiatric disorders 精神疾患モデルとしてのGADノックアウトラット Fujihara Kazuyuki（藤原 和之），柳川 右千夫 Department of Genetic and Behavioral Neuroscience, Graduate School of Medicine, Gunma University. Cognitive impairment including working memory (WM) deficit in schizophrenia is an important predictor of functional outcome of the patients. Reduction of glutamate decarboxylase 67 (GAD67) seen in the postmortem brain of schizophrenia is suggested to be a key pathophysiological basis of WM deficit. Since GAD67 is one of the enzymes which produce inhibitory neurotransmitter GABA, it has an important role to maintain adequate excitatory-inhibitory balance in the neuronal circuit. Unfortunately, a conventional GAD67 knockout mouse is not useful as a model animal of WM impairment of schizophrenia, because they are mortal at the first day after birth. Recently, several distinct GAD67 conditional knockout/knockdown mice have been reported, and some of them show behavioral alterations relevant to schizophrenia. But WM alteration in these model mice has not been demonstrated so far. Here, we will show that GAD67 knockout (GAD67-/-) rats, newly generated by CRISPER/Cas9 genome editing, exhibit WM impairment. Surprisingly, at least half of GAD67-/- rats can survive to adulthood not as GAD67-/- mice. We confirmed complete deletion of GAD67 protein in adult brain of GAD67-/- rats by western blot analysis and immunohistochemistry. GAD67-/- rats actually show lower performance in the Morris water maze task and the 8-arm radial maze task compared to GAD67+/+ littermates. These results suggest that the GAD67 loss leads to impairments in spatial learning and WM. As far as we know, this is the first direct evidence that GAD67 reduction causes WM impairment. GAD-/- rats will be a promised candidate for model animal of cognitive impairment of schizophrenia. SY8-3 GABA synthesis in mouse models of fear- and stress-related psychopathology 恐怖およびストレスに関連した精神障害を示すマウスモデルにおけるGABA合成について Stork Oliver Department of Genetics & Molecular Neurobiology, Institute of Biology, Otto-von-Guericke University Magdeburg, and Center of Behavioral Brain Sciences Pavlovian fear conditioning is an established learning paradigm that allows studying neural mechanisms of fear and anxiety in various species and that may be employed to emulate specific aspects of anxiety disorders. We investigated, in mice, molecular and physiological processes in the amygdalo-hippocampal system that are involved in the consolidation and reconsolidation of such long-term fear memory. GABAergic interneurons are of critical importance for these processes, as indicated by the hyperarousal, fear generalization and deficit in fear extinction of mice deficient for the key enzyme in GABA synthesis, glutamic acid decarboxylase 65. We observed specific changes in the expression of glutamic acid decarboxylase and other interneuron-specific factors in hippocampal subfields following the acquisition and reconsolidation of fear, and in response to the enhancement of fear conditioning in a juvenile stress model of PTSD. The observed changes in gene expression could be attributed to specific subpopulations of GABAergic interneurons and their role in stress- and fear memory-induced alterations of local circuit activity of dentate gyrus, area CA3 and area CA1. Targeted genetic and chemogenetic manipulation of these interneuron populations resulted in alterations of spontaneous and induced network activity, including sharp wave ripples and gamma oscillations, that were associated with fear memory generalization and lack of fear extinction in the behaving animals. Together, these data demonstrate the role of GABAergic interneurons in the hippocampus in the development of pathological fear memory and its physiological network correlates. SY8-4 Prenatal stress to the GAD67 heterozygous mice impairs GABA neurogenesis followed by physiological dysfunctions in postnatal life GAD67ヘテロ接合体マウスへの胎生期ストレス負荷がもたらすGABAニューロン発生障害とシナプス伝達・脳波・行動の異常 Fukuda Atsuo（福田 敦夫） Dept. of Neurophysiol., Hamamatsu Univ. Sch. of Med. Exposure to prenatal stress (PS) and mutations in Gad1 encodig the GABA synthesizing enzyme glutamate decarboxylase (GAD) 67, are both risk factors for psychiatric disorders. Abnormalities of parvalbumin (PV)-positive GABAergic interneurons in the medial prefrontal cortex (mPFC) and hippocampus (HIP) are often observed in schizophrenia and autistic patients. To elucidate their relationship, we examined GAD67-GFP knock-in mice (GAD67 +/GFP ) that underwent PS. BrdU-assessed neurogenesis of GABAergic neurons was significantly diminished, and postnatally, the density of PV-positive but not PV-negative GABAergic neurons was significantly decreased in the mPFC, HIP and somatosensory cortex. Neither wild type (WT) litter mates underwent PS nor naive GAD67+/GFP offspring showed these findings, suggesting that PS plus Gad1 anomaly could specifically disturb the proliferation of PV-positive neurons. Behavioral comparisons (WT-PS, WT-naive, GAD67+/GFP-PS, GAD67+/GFP-naive) indicated abnormal social interaction and pre-pulse inhibition only in GAD67+/GFP-PS, supporting gene-environment interaction. We found multiple genes related to neurogenesis and/or neural migration were hyper- or hypo-methylated in GAD67+/GFP-PS and some were associated with expressional alterations. Also, expression of fukutin, which glycosylates α-dystroglycan (α-DG), was suppressed. Glycosylation of α-DG, that promotes plasticity of GABAergic synapses, decreased. So we examined inhibitory postsynaptic currents and tonic inhibition by spilt-over GABA, and found they were significantly altered in mPFC. Electrocorticogram showed reduction in γ-frequency mean power spectrum density in mPFC. These findings suggest new insights into underlying GABAergic mechanisms of the pathology of psychiatric disorders. SY8-5 Postmortem brain analysis on cortical E-I balance in cognitive impairments in schizophrenia 死後脳研究から考える統合失調症の認知機能障害とE-I balance変化の分子基盤 Kimoto Sohei（紀本 創兵） Department of Psychiatry, Nara Medical University School of Medicine, Kashihara, Japan Impairments in certain cognitive function that reflect dysfunction of dorsolateral prefrontal cortex (DLPFC) circuitry are, in a greater or lesser extent, observed in schizophrenia. Disturbed excitation and inhibition balance (E/I balance) in neuronal circuits have been implicated in cognitive impairments of schizophrenia. Given the role of activity-dependent transcription in synapse plasticity, disease-related alterations in activity-dependent gene expression might contribute to the molecular basis of cognitive impairments in the illnesses. Here, using postmortem brain samples, we examined levels of the activity-regulated immediate early genes (IEGs) in the DLPFC of schizophrenia and healthy comparison subjects. In schizophrenia subjects, mRNA levels of neuronal activity-regulated pentraxins 2 (NARP), which can regulate excitatory inputs from pyramidal neurons to parvalbumin-containing interneurons (PV-INs), were lower in the DLPFC pyramidal cells, suggesting that lower activity in excitatory neurons might lead to less NARP expression which in turn results in weaker excitatory drive into PV-Ins in schizophrenia. In addition, mRNA levels of the transcriptional regulatory factor Zif268, which can regulate glutamic acid decarboxylase 67-kDa (GAD67) expression, were lower in the DLPFC PV-INs in schizophrenia subjects, suggesting that reduced transcription of Zif268 might mediate the lower levels of GAD67 expression observed in schizophrenia subjects. These findings were robust to the effects of the confounding variables examined and differed from other IEGs. We suggest lower levels of NARP and ZIf268 transcripts in the DLPFC might contribute to the molecular substrate for disturbed E/I balance, providing a new insight into cognitive impairment in schizophrenia.